Novel Strategies for Target Design and Gene Silencing in the Insect Vectors of Plant Pathogens

RNA interference (RNAi) has become the new paradigm for testing the key gene regulatory pathways in insect vectors of plant pathogens. The key molecules at the center of this multi-step disruptive process are double stranded RNAs (dsRNAs) that are known to be triggers for siRNA biogenesis. A robust systemic RNAi, though elegantly present in C. elegans, is absent in hemipteran insect vectors largely due to the absence of RNA dependent RNA polymerase (RdRP). Due to this issue, there is a need for identification of effective RNAi targets and for the development of efficient delivery vehicles. The recent advances in Next Generation Sequencing technologies allow us to generate small RNA profiles in insect vectors that display a robust silencing signals against the invading foreign organisms. In our experiments with the glassy-winged sharpshooter (Homalodisca vitripennis) and the Asian citrus psyllid (Diaphorina citri), we have identified small RNAs against viruses that naturally infect these vectors. Further, these new technologies (Transcriptome/Genome sequencing) enabled us to identify gene targets for specific activity by RNAi. We recently began novel experiments to deliver the dsRNAs and artificial microRNAs through the use of viral vectors to effectively silence insect RNAi targets. These transient approaches combined with spatial delivery of hairpin RNAs in transgenic plants are two of the strategies that we employ for gene silencing in hemipteran insect pests.